The overall objective of this proposal is to demonstrate that laser pre-illumination can enhance skin-to-liver delivery of sporozoites (SPZs), augmenting SPZ-based vaccines. An effective, licensed anti-malarial vaccine remains an urgent need for prevention against plasmodium infection. To date, the gold standard for malarial vaccine developers is still immunization by bites of irradiation (g), SPZ-infected mosquitoes. However, translation of this approach into the clinic faces formidable obstacles, due to impracticality of vaccinating a large number of people by bites of infectious mosquitoes and unethic of this route of immunization. For SPZ- based vaccines, irrespective of whether the SPZs are attenuated by irradiation or genetic modification, the efficacy is correlated with the number of SPZs trafficking into the liver. While intravenous (IV) injection may be most sufficient in delivering SPZs to the liver, it is not a clinically approved route of immunization. On the othe hand, intradermal (ID) vaccination, not only mimicking natural infection but also a clinically acceptable route, is much less efficient than IV because emigration of SPZs out of the skin is restricted by the densely packed network in the dermis, which may be particularly true for cryopreserved gSPZs owing to their reduced motility. We found that laser pre-illumination of the inoculation site prior to ID enhanced skin-to-liver traffic of gSPZs to a level comparable to IV injection. The illumination transiently disrupted the dense microarchitecture of the skin, permitting relatively free movement of SPZs and their entry into the capillaries and lymphatic vessels in the dermis. We hypothesize that this laser treatment can augment ID delivery of SPZ-based vaccines and when combined with a-GalCer (a-galactosylceramide), an adjuvant for SPZ-induced immunity, can further boost SPZ-induced protection against malaria. To test this hypothesis, we will verify whether laser-mediated facilitation of skin-to-liver delivery of SPZs is SPZ dose- and/or motility-dependent by quantification of the number of SPZs in the liver 40 hr after ID injection of varying numbers of SPZs into the site of laser illumination. This dose-dependent effect will be then tested with freshly isolated or cryopreserved gSPZs or genetically attenuated SPZs. We will also validate in outbred and inbred mouse models that laser illumination followed by ID inoculation of cryopreserved gSPZs or genetically attenuated SPZs affords similar protection against virulent SPZ infections as IV vaccination and a synergistic effect between laser and a-GalCer adjuvant on the protective immunity. The preclinical study, if successful, can potentially take SPZ-based vaccines one step closer to their clinical application.